PROJECT SUMMARY Type 1 diabetes (T1D) is a chronic autoimmune disorder that affects ~1% of population worldwide. Exogenous insulin treatment is the standard of care for T1D, but often negatively affects the quality of life and is ineffective in preventing recurrent hyperglycemia episodes and chronic complications. Recent studies show that human islet allografts can restore long-term normoglycemia and insulin independence, protect from severe hypoglycemia, and slow progression of microvascular lesions in immunosuppressed T1D patients. However, immune rejection and continuous use of immunosuppression to control rejection are two major limitations of clinical islet transplantation. Standard immunosuppression is ineffective in achieving long-term graft survival and also has significant adverse effects on the graft and graft recipients. Therefore, the development of novel approaches to prevent rejection of islet grafts without chronic immunosuppression is a significant goal. Allogeneic islets are subject to rejection by both alloreactive and autoreactive T effector (Teff) cells. An imbalance in the frequency of pathogenic Teff and protective T regulatory (Treg) cells is the underlying cause of T1D and allogeneic islet graft rejection. Restoring the physiological Teff and Treg balance has significant therapeutic potential. Approaches attempting to tilt the balance in favor of Treg cells have so far targeted either Teff or Treg cells individually for modulation with limited success. The primary goal of this application is to target both cell types simultaneously for modulation for an outcome in favor of Treg cell expansion. This will be achieved using innovative polyethylene glycol hydrogel particle platforms for graft-targeted delivery and controlled presentation of two novel biologics serving as agonists of Fas and IL-2R receptors. Teff cells activated by antigens express Fas receptor and become sensitive to FasL-mediated apoptosis. IL-2R signaling preferentially sensitizes Teff cells to Fas-induced apoptosis and is also required for Treg cells (CD4+CD25+FoxP3+) generation, expansion, and survival. Therefore, we hypothesize that the combined use of agonists of Fas and IL-2R will preferentially eliminate Teff cells and generate/expand Treg cells within the graft microenvironment, resulting in induced-immune privilege and sustained survival and function of islet allograft in the absence of any immunosuppression. A set of preliminary data support this hypothesis and provide strong scientific premise and feasibility for this application. This concept will be tested in three different allogeneic islet transplantation settings for efficacy and mechanisms; chemically diabetic BALB/c-to- C57BL/6 mice, spontaneously diabetic C57BL/6-to-NOD mice, and human islets into humanized mice. These models will generate critical data relevant to the human setting. Furthermore, proof-of-efficacy and the elucidation of the immune mechanisms regulating effective outcomes will expedite further refinement of this immunomodulatory concept and its eventual translation to nonhuman primates as a prelude to clinical trials for the treatment of type 1 diabetes.